The covalent attachment of one or multiple heme cofactors to cytochrome c protein chains enables cytochrome c proteins to be used in electron transfer and redox catalysis in extracytoplasmic environments. A dedicated heme maturation machinery, whose core component is a heme lyase, scans nascent peptides after Sec-dependent translocation for CXnCH-binding motifs. Here we report the three-dimensional (3D) structure of the heme lyase CcmF, a 643-amino acid integral membrane protein, from Thermus thermophilus. CcmF contains a heme b cofactor at the bottom of a large cavity that opens toward the extracellular side to receive heme groups from the heme chaperone CcmE for cytochrome maturation. A surface groove on CcmF may guide the extended apoprotein to heme attachment at or near a loop containing the functionally essential WXWD motif, which is situated above the putative cofactor binding pocket. The structure suggests heme delivery from within the membrane, redefining the role of the chaperone CcmE.Deciphering mechanisms in cell-fate decisions requires single-cell holistic reconstructions of multidimensional epigenomic states in transcriptional regulation. Here we develop CoTECH, a combinatorial barcoding method allowing high-throughput single-cell joint detection of chromatin occupancy and transcriptome. We used CoTECH to examine bivalent histone marks (H3K4me3 and H3K27me3) with transcription from naive to primed mouse embryonic stem cells. https://www.selleckchem.com/products/BIRB-796-(Doramapimod).html We also derived concurrent bivalent marks in pseudosingle cells using transcriptome as an anchor for resolving pseudotemporal bivalency trajectories and disentangling a context-specific interplay between H3K4me3/H3K27me3 and transcription level. Next, we revealed the regulatory basis of endothelial-to-hematopoietic transition in two waves of hematopoietic cells and distinctive enhancer-gene-linking schemes guiding hemogenic endothelial cell emergence, indicating a unique epigenetic control of transcriptional regulation for hematopoietic stem cell priming. CoTECH provides an efficient framework for single-cell coassay of chromatin occupancy and transcription, thus enabling higher-dimensional epigenomic reconstructions.The biological roles of DNA methylation have been elucidated by profiling methods based on whole-genome or reduced-representation bisulfite sequencing, but these approaches do not efficiently survey the vast numbers of non-coding regulatory elements in mammalian genomes. Here we present an extended-representation bisulfite sequencing (XRBS) method for targeted profiling of DNA methylation. Our design strikes a balance between expanding coverage of regulatory elements and reproducibly enriching informative CpG dinucleotides in promoters, enhancers and CTCF binding sites. Barcoded DNA fragments are pooled before bisulfite conversion, allowing multiplex processing and technical consistency in low-input samples. Application of XRBS to single leukemia cells enabled us to evaluate genetic copy number variations and methylation variability across individual cells. Our analysis highlights heterochromatic H3K9me3 regions as having the highest cell-to-cell variability in their methylation, likely reflecting inherent epigenetic instability of these late-replicating regions, compounded by differences in cell cycle stages among sampled cells.Continuous supply of immune cells throughout life relies on the delicate balance in the hematopoietic stem cell (HSC) pool between long-term maintenance and meeting the demands of both normal blood production and unexpected stress conditions. Here we identified distinct subsets of human long-term (LT)-HSCs that responded differently to regeneration-mediated stress an immune checkpoint ligand CD112lo subset that exhibited a transient engraftment restraint (termed latency) before contributing to hematopoietic reconstitution and a primed CD112hi subset that responded rapidly. This functional heterogeneity and CD112 expression are regulated by INKA1 through direct interaction with PAK4 and SIRT1, inducing epigenetic changes and defining an alternative state of LT-HSC quiescence that serves to preserve self-renewal and regenerative capacity upon regeneration-mediated stress. Collectively, our data uncovered the molecular intricacies underlying HSC heterogeneity and self-renewal regulation and point to latency as an orchestrated physiological response that balances blood cell demands with preserving a stem cell reservoir.A key driver of patients' well-being and clinical trials for Parkinson's disease (PD) is the course that the disease takes over time (progression and prognosis). To assess how genetic variation influences the progression of PD over time to dementia, a major determinant for quality of life, we performed a longitudinal genome-wide survival study of 11.2 million variants in 3,821 patients with PD over 31,053 visits. We discover RIMS2 as a progression locus and confirm this in a replicate population (hazard ratio (HR)?=?4.77, P?=?2.78?×?10-11), identify suggestive evidence for TMEM108 (HR?=?2.86, P?=?2.09?×?10-8) and WWOX (HR?=?2.12, P?=?2.37?×?10-8) as progression loci, and confirm associations for GBA (HR?=?1.93, P?=?0.0002) and APOE (HR?=?1.48, P?=?0.001). Polygenic progression scores exhibit a substantial aggregate association with dementia risk, while polygenic susceptibility scores are not predictive. This study identifies a novel synaptic locus and polygenic score for cognitive disease progression in PD and proposes diverging genetic architectures of progression and susceptibility.Animal genomes are partitioned and folded at various scales that contribute distinctly to nuclear processes. While structural features have been disrupted either globally or at select loci in loss-of-function studies, gain-of-function studies that probe the role of genome architecture have lagged behind. Here we examine recent advances in experimentally creating chromatin loops, contact domains, boundaries and compartments. Furthermore, we explore parallels between this emerging theme and natural evolution of mammalian genomes with increasing architectural complexity. Finally, we provide a perspective on how insights arising from recent gain-of-function studies may inform future endeavors toward engineering the three-dimensional genome.